Transcriptomic analysis of seed development in Paysonia auriculata (Brassicaceae) identifies genes involved in hydroxy fatty acid biosynthesis.

Paysonia auriculata (Brassicaceae) produces multiple hydroxy fatty acids as major components of the seed oil. We tracked the changes in seed oil composition and gene expression during development, starting 14 days after flowers had been pollinated. Seed oil changes showed initially higher levels of saturated and unsaturated fatty acids (FAs) but little accumulation of hydroxy fatty acids (HFAs). Starting 21 days after pollination (DAP) HFA content sharply increased, and reached almost 30% at 28 DAP. Total seed oil also increased from a low of approximately 2% at 14 DAP to a high of approximately 20% by 42 DAP. We identified almost all of the fatty acid synthesis and modification genes that are known from Arabidopsis, and, in addition, a strong candidate for the hydroxylase gene that mediates the hydroxylation of fatty acids to produce valuable hydroxy fatty acids (HFAs) in this species. The gene expression network revealed is very similar to that of the emerging oil crop, Physaria fendleri, in the sister genus to Paysonia. Phylogenetic analyses indicate the hydroxylase enzyme, FAH12, evolved only once in Paysonia and Physaria, and that the enzyme is closely related to FAD2 enzymes. Phylogenetic analyses of FAD2 and FAH12 in the Brassicaceae and outgroup genera suggest that the branch leading to the hydroxylase clade of Paysonia and Physaria is under relaxed selection, compared with the strong purifying selection found across the FAD2 lineages.

Functional Evaluation of Splicing for Variants of Uncertain Significance in Patients with Inherited Retinal Diseases.

Inherited retinal diseases (IRD) comprise a heterogeneous set of clinical and genetic disorders that lead to blindness. Given the emerging opportunities in precision medicine and gene therapy, it has become increasingly important to determine whether DNA variants with uncertain significance (VUS) are responsible for patients' IRD. This research was performed to assess the functional consequence of six VUS identified in patients with IRD. Clinical assessments included an ophthalmic examination, best-corrected visual acuity, and kinetic perimetry. Imaging was acquired with the Optos ultra-widefield camera and spectral domain optical coherence tomography (SD-OCT). Genetic testing was performed by Molecular Vision Laboratories. VUS that were predicted to alter splicing were analyzed with a minigene assay, which revealed that VUS in the genes OPA1, CNGB1, and CLUAP1 altered spicing mechanisms. Due to emerging gene and cell therapies, these results expand the genotype-phenotype correlations for patients diagnosed with an IRD.

Integration of high-density genetic mapping with transcriptome analysis uncovers numerous agronomic QTL and reveals candidate genes for the control of tillering in sorghum.

Phenotypes such as branching, photoperiod sensitivity, and height were modified during plant domestication and crop improvement. Here, we perform quantitative trait locus (QTL) mapping of these and other agronomic traits in a recombinant inbred line (RIL) population derived from an interspecific cross between Sorghum propinquum and Sorghum bicolor inbred Tx7000. Using low-coverage Illumina sequencing and a bin-mapping approach, we generated ∼1920 bin markers spanning ∼875 cM. Phenotyping data were collected and analyzed from two field locations and one greenhouse experiment for six agronomic traits, thereby identifying a total of 30 QTL. Many of these QTL were penetrant across environments and co-mapped with major QTL identified in other studies. Other QTL uncovered new genomic regions associated with these traits, and some of these were environment-specific in their action. To further dissect the genetic underpinnings of tillering, we complemented QTL analysis with transcriptomics, identifying 6189 genes that were differentially expressed during tiller bud elongation. We identified genes such as Dormancy Associated Protein 1 (DRM1) in addition to various transcription factors that are differentially expressed in comparisons of dormant to elongating tiller buds and lie within tillering QTL, suggesting that these genes are key regulators of tiller elongation in sorghum. Our study demonstrates the usefulness of this RIL population in detecting domestication and improvement-associated genes in sorghum, thus providing a valuable resource for genetic investigation and improvement to the sorghum community.

Variable expressivity in patients with autosomal recessive retinitis pigmentosa associated with the gene CNGB1.

PURPOSE: In a cohort of eight families (11 patients) with autosomal recessive retinitis pigmentosa (arRP), we clinically characterized disease associated with mutations in CNGB1. METHODS: Visual function was determined by measuring the patients' visual acuity, dark- and light-adapted perimetry, and by full-field electroretinography. Retinal structure was evaluated with spectral-domain optical coherence tomography, fundus imaging, and autofluorescence imaging. RESULTS: Age of onset ranged from 4 to 49 years (mean [SD] 26 [17], median 27 years). The age at visit was 27-54 years, mean 37 (17). The range of visual acuity was logMAR -0.1 to 1.3 (Snellen 20/16 to 20/400) in the right eye and -0.1 to 0.9 (Snellen 20/16 to 20/160) in the left eye. Electrophysiological testing in five patients showed an absence of the rod response. Cone responses ranged from normal to severely reduced. The patients exhibited loss of rod vision more severe than cone vision. Funduscopic images showed widespread retinal degeneration with pigment clumping, optic disk pallor, arteriole attenuation, and a peri-foveal ring of hyper autofluorescence. Three families were tested for olfactory dysfunction and results indicated mild to complete anosmia in individuals with mutations in CNGB1. Genetic analysis revealed 6 novel variants, c.2127 C > G, p.Phe709Leu; c.1431 C > A, p.Cys477*; c.2034 G > A, p.Trp678*; c.2092 T > C, p.Cys698Arg; and c.583 + 2 T > C, c.2305-34 G > A and 3 variants that have been previously described, c.2957A>T, p.Asn986Ile; c.2544dup, p.Leu849Alafs*3; and c.2492 + 1 G > A. DISCUSSION: This is the first report for six novel CNGB1 variants associated with arRP. Two families had olfactory dysfunction in patients with arRP and family members who were heterozygous for a CNGB1 mutation. Additionally, findings demonstrated variable penetrance and expressivity of disease in these patients.

Domestication and Improvement in the Model C4 Grass, Setaria.

Setaria viridis (green foxtail) and its domesticated relative S. italica (foxtail millet) are diploid C4 panicoid grasses that are being developed as model systems for studying grass genomics, genetics, development, and evolution. According to archeological evidence, foxtail millet was domesticated from green foxtail approximately 9,000 to 6,000 YBP in China. Under long-term human selection, domesticated foxtail millet developed many traits adapted to human cultivation and agricultural production. In comparison with its wild ancestor, foxtail millet has fewer vegetative branches, reduced grain shattering, delayed flowering time and less photoperiod sensitivity. Foxtail millet is the only present-day crop in the genus Setaria, although archeological records suggest that other species were domesticated and later abandoned in the last 10,000 years. We present an overview of domestication in foxtail millet, by reviewing recent studies on the genetic regulation of several domesticated traits in foxtail millet and discuss how the foxtail millet and green foxtail system could be further developed to both better understand its domestication history, and to provide more tools for future breeding efforts.

The C4 Model Grass Setaria Is a Short Day Plant with Secondary Long Day Genetic Regulation.

The effect of photoperiod (day:night ratio) on flowering time was investigated in the wild species, Setaria viridis, and in a set of recombinant inbred lines (RILs) derived from a cross between foxtail millet (S. italica) and its wild ancestor green foxtail (S. viridis). Photoperiods totaled 24 h, with three trials of 8:16, 12:12 and 16:8 light:dark hour regimes for the RIL population, and these plus 10:14 and 14:10 for the experiments with S. viridis alone. The response of S. viridis to light intensity as well as photoperiod was assessed by duplicating photoperiods at two light intensities (300 and 600 µmol.m-2.s-1). In general, day lengths longer than 12 h delayed flowering time, although flowering time was also delayed in shorter day-lengths relative to the 12 h trial, even when daily flux in high intensity conditions exceeded that of the low intensity 12 h trial. Cluster analysis showed that the effect of photoperiod on flowering time differed between sets of RILs, with some being almost photoperiod insensitive and others being delayed with respect to the population as a whole in either short (8 or 12 h light) or long (16 h light) photoperiods. QTL results reveal a similar picture, with several major QTL colocalizing between the 8 and 12 h light trials, but with a partially different set of QTL identified in the 16 h trial. Major candidate genes for these QTL include several members of the PEBP protein family that includes Flowering Locus T (FT) homologs such as OsHd3a, OsRFT1, and ZCN8/12. Thus, Setaria is a short day plant (flowering quickest in short day conditions) whose flowering is delayed by long day lengths in a manner consistent with the responses of most other members of the grass family. However, the QTL results suggest that flowering time under long day conditions uses additional genetic pathways to those used under short day conditions.

Development and Genetic Control of Plant Architecture and Biomass in the Panicoid Grass, Setaria.

The architecture of a plant affects its ability to compete for light and to respond to environmental stresses, thus affecting overall fitness and productivity. Two components of architecture, branching and height, were studied in 182 F7 recombinant inbred lines (RILs) at the vegetative, flowering and mature developmental stages in the panicoid C4 model grass system, Setaria. The RIL population was derived from a cross between domesticated S. italica (foxtail millet) and its wild relative S. viridis (green foxtail). In both field and greenhouse trials the wild parent was taller initially, started branching earlier, and flowered earlier, while the domesticated parent was shorter initially, but flowered later, producing a robust tall plant architecture with more nodes and leaves on the main culm and few or no branches. Biomass was highly correlated with height of the plant and number of nodes on the main culm, and generally showed a negative relationship with branch number. However, several of the RILs with the highest biomass in both trials were significantly more branched than the domesticated parent of the cross. Quantitative trait loci (QTL) analyses indicate that both height and branching are controlled by multiple genetic regions, often with QTL for both traits colocalizing in the same genomic regions. Genomic positions of several QTL colocalize with QTL in syntenic regions in other species and contain genes known to control branching and height in sorghum, maize, and switchgrass. Included in these is the ortholog of the rice SD-1 semi-dwarfing gene, which underlies one of the major Setaria height QTL. Understanding the relationships between height and branching patterns in Setaria, and their genetic control, is an important step to gaining a comprehensive knowledge of the development and genetic regulation of panicoid grass architecture.

Morphological diversity and genetic regulation of inflorescence abscission zones in grasses.

UNLABELLED: • PREMISE OF THE STUDY: Variation in how seeds are dispersed in grasses is ecologically important, and selection for dispersal mechanisms has produced a great variety of dispersal structures (diaspores). Abscission ("shattering") is necessary in wild grasses, but its elimination by selection on nonshattering mutants was a key component of the domestication syndrome in cereal grasses. A key question is whether a common genetic pathway controls abscission in wild grasses, and, if so, what genes in that pathway may have been selected upon during domestication. We summarize morphological and genetic information on abscission zones and disarticulation patterns in grasses and identify hypotheses to test the likelihood of a common genetic pathway.• METHODS: Morphological data on abscission zones for over 10000 species of grasses were tabulated and analyzed using a tribal phylogeny of the grasses. The genomic location of quantitative trait loci (QTLs) and orthologs of genes controlling shattering were compared across species to ascertain whether the same loci might control shattering in different grass lineages.• RESULTS AND CONCLUSIONS: The simple trait of nonshattering is derived from a great diversity of shattering phenotypes. Several sets of QTLs from multiple species are syntenic yet many are not. Genes known to be involved in shattering in several species were found to have orthologs that sometimes colocalized with QTLs in different species, adding support to the hypothesis of retention of a common genetic pathway. These results are used to suggest a research plan that could test the common genetic pathway model more thoroughly.

Beyond the single gene: How epistasis and gene-by-environment effects influence crop domestication.

Domestication is a multifaceted evolutionary process, involving changes in individual genes, genetic interactions, and emergent phenotypes. There has been extensive discussion of the phenotypic characteristics of plant domestication, and recent research has started to identify the specific genes and mutational mechanisms that control domestication traits. However, there is an apparent disconnect between the simple genetic architecture described for many crop domestication traits, which should facilitate rapid phenotypic change under selection, and the slow rate of change reported from the archeobotanical record. A possible explanation involves the middle ground between individual genetic changes and their expression during development, where gene-by-gene (epistatic) and gene-by-environment interactions can modify the expression of phenotypes and opportunities for selection. These aspects of genetic architecture have the potential to significantly slow the speed of phenotypic evolution during crop domestication and improvement. Here we examine whether epistatic and gene-by-environment interactions have shaped how domestication traits have evolved. We review available evidence from the literature, and we analyze two domestication-related traits, shattering and flowering time, in a mapping population derived from a cross between domesticated foxtail millet and its wild progenitor. We find that compared with wild progenitor alleles, those favored during domestication often have large phenotypic effects and are relatively insensitive to genetic background and environmental effects. Consistent selection should thus be able to rapidly change traits during domestication. We conclude that if phenotypic evolution was slow during crop domestication, this is more likely due to cultural or historical factors than epistatic or environmental constraints.

Genetic control and comparative genomic analysis of flowering time in Setaria (Poaceae).

We report the first study on the genetic control of flowering in Setaria, a panicoid grass closely related to switchgrass, and in the same subfamily as maize and sorghum. A recombinant inbred line mapping population derived from a cross between domesticated Setaria italica (foxtail millet) and its wild relative Setaria viridis (green millet), was grown in eight trials with varying environmental conditions to identify a small number of quantitative trait loci (QTL) that control differences in flowering time. Many of the QTL across trials colocalize, suggesting that the genetic control of flowering in Setaria is robust across a range of photoperiod and other environmental factors. A detailed comparison of QTL for flowering in Setaria, sorghum, and maize indicates that several of the major QTL regions identified in maize and sorghum are syntenic orthologs with Setaria QTL, although the maize large effect QTL on chromosome 10 is not. Several Setaria QTL intervals had multiple LOD peaks and were composed of multiple syntenic blocks, suggesting that observed QTL represent multiple tightly linked loci. Candidate genes from flowering time pathways identified in rice and Arabidopsis were identified in Setaria QTL intervals, including those involved in the CONSTANS photoperiod pathway. However, only three of the approximately seven genes cloned for flowering time in maize colocalized with Setaria QTL. This suggests that variation in flowering time in separate grass lineages is controlled by a combination of conserved and lineage specific genes.

Reference genome sequence of the model plant Setaria.

We generated a high-quality reference genome sequence for foxtail millet (Setaria italica). The ∼400-Mb assembly covers ∼80% of the genome and >95% of the gene space. The assembly was anchored to a 992-locus genetic map and was annotated by comparison with >1.3 million expressed sequence tag reads. We produced more than 580 million RNA-Seq reads to facilitate expression analyses. We also sequenced Setaria viridis, the ancestral wild relative of S. italica, and identified regions of differential single-nucleotide polymorphism density, distribution of transposable elements, small RNA content, chromosomal rearrangement and segregation distortion. The genus Setaria includes natural and cultivated species that demonstrate a wide capacity for adaptation. The genetic basis of this adaptation was investigated by comparing five sequenced grass genomes. We also used the diploid Setaria genome to evaluate the ongoing genome assembly of a related polyploid, switchgrass (Panicum virgatum).

Phylogenetic analysis of seven WRKY genes across the palm subtribe Attaleinae (Arecaceae) [corrected] identifies Syagrus as sister group of the coconut.

BACKGROUND: The Cocoseae is one of 13 tribes of Arecaceae subfam. Arecoideae, and contains a number of palms with significant economic importance, including the monotypic and pantropical Cocos nucifera L., the coconut, the origins of which have been one of the "abominable mysteries" of palm systematics for decades. Previous studies with predominantly plastid genes weakly supported American ancestry for the coconut but ambiguous sister relationships. In this paper, we use multiple single copy nuclear loci to address the phylogeny of the Cocoseae subtribe Attaleinae, and resolve the closest extant relative of the coconut. METHODOLOGY/PRINCIPAL FINDINGS: We present the results of combined analysis of DNA sequences of seven WRKY transcription factor loci across 72 samples of Arecaceae tribe Cocoseae subtribe Attaleinae, representing all genera classified within the subtribe, and three outgroup taxa with maximum parsimony, maximum likelihood, and Bayesian approaches, producing highly congruent and well-resolved trees that robustly identify the genus Syagrus as sister to Cocos and resolve novel and well-supported relationships among the other genera of the Attaleinae. We also address incongruence among the gene trees with gene tree reconciliation analysis, and assign estimated ages to the nodes of our tree. CONCLUSIONS/SIGNIFICANCE: This study represents the as yet most extensive phylogenetic analyses of Cocoseae subtribe Attaleinae. We present a well-resolved and supported phylogeny of the subtribe that robustly indicates a sister relationship between Cocos and Syagrus. This is not only of biogeographic interest, but will also open fruitful avenues of inquiry regarding evolution of functional genes useful for crop improvement. Establishment of two major clades of American Attaleinae occurred in the Oligocene (ca. 37 MYBP) in Eastern Brazil. The divergence of Cocos from Syagrus is estimated at 35 MYBP. The biogeographic and morphological congruence that we see for clades resolved in the Attaleinae suggests that WRKY loci are informative markers for investigating the phylogenetic relationships of the palm family.